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Sandra Denery-papini - One of the best experts on this subject based on the ideXlab platform.
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Deamidation and enzymatic hydrolysis of Gliadins alter their processing by dendritic cells in vitro
Journal of Agricultural and Food Chemistry, 2020Co-Authors: Clélia Villemin, Sandra Denery-papini, Véronique Solé-jamault, Olivier Tranquet, Joost J. Smit, Raymond H. H. Pieters, Grégory BouchaudAbstract:Gliadins are major wheat allergens. Their treatment by acid or enzymatic hydrolysis has been shown to modify their allergenic potential. As the interaction of food proteins with dendritic cells (DCs) is a key event in allergic sensitization, we wished to investigate whether deamidation and enzymatic hydrolysis influence Gliadin processing by DC and to examine the capacity of Gliadins to activate DCs. We compared the uptake and degradation of native and modified Gliadins by DCs using mouse bone marrow-derived DCs. We also analyzed the effects of these interactions on the phenotypes of DCs and T helper (Th) lymphocytes. Modifying Gliadins induced a change in physicochemical properties (molecular weight, hydrophobicity, and sequence) and also in the peptide size. These alterations in turn led to increased uptake and intracellular degradation of the proteins by DCs. Native Gliadins (NGs) (100 μg/mL), but not modified Gliadins, increased the frequency of DC expressing CD80 (15.41 ± 2.36% vs 6.81 ± 1.10%, p < 0.001), CCR7 (28.53 ± 8.17% vs 17.88 ± 2.53%, p < 0.001), CXCR4 (70.14 ± 4.63% vs 42.82 ± 1.96%, p < 0.001), and CCR7-dependent migration (2.46 ± 1.45 vs 1.00 ± 0.22, p < 0.01) compared with NGs. This was accompanied by Th lymphocyte activation (30.37 ± 3.87% vs 21.53 ± 3.14%, p < 0.1) and proliferation (16.39 ± 3.97% vs 9.31 ± 2.80%, p > 0.1). Moreover, hydrolysis decreases the peptide size and induces an increase in Gliadin uptake and degradation. Deamidation and extensive enzymatic hydrolysis of Gliadins modify their interaction with DCs, leading to alteration of their immunostimulatory capacity. These findings demonstrate the strong relationship between the biochemical characteristics of proteins and immune cell interactions.
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Towards reducing the immunogenic potential of wheat flour: omega Gliadins encoded by the D genome of hexaploid wheat may also harbor epitopes for the serious food allergy WDEIA
BMC Plant Biology, 2018Co-Authors: Susan B. Altenbach, Sandra Denery-papini, You-ran Jang, Annamaria Simon-buss, Sun-hyung Lim, Naxin Huo, Han-chang Chang, Florence Pineau, Chon-sik Kang, Jong-yeol LeeAbstract:Background: Omega-5 Gliadins are a group of highly repetitive gluten proteins in wheat flour encoded on the 1B chromosome of hexaploid wheat. These proteins are the major sensitizing allergens in a severe form of food allergy called wheat-dependent exercise-induced anaphylaxis (WDEIA). The elimination of omega-5 Gliadins from wheat flour through biotechnology or breeding approaches could reduce the immunogenic potential and adverse health effects of the flour. Results: A mutant line missing low-molecular weight glutenin subunits encoded at the Glu-B3 locus was selected previously from a doubled haploid population generated from two Korean wheat cultivars. Analysis of flour from the mutant line by 2-dimensional gel electrophoresis coupled with tandem mass spectrometry revealed that the omega-5 Gliadins and several gamma Gliadins encoded by the closely linked Gli-B1 locus were also missing as a result of a deletion of at least 5.8Mb of chromosome 1B. Two-dimensional immunoblot analysis of flour proteins using sera from WDEIA patients showed reduced IgE reactivity in the mutant relative to the parental lines due to the absence of the major omega-5 Gliadins. However, two minor proteins showed strong reactivity to patient sera in both the parental and the mutant lines and also reacted with a monoclonal antibody against omega-5 Gliadin. Analysis of the two minor reactive proteins by mass spectrometry revealed that both proteins correspond to omega-5 Gliadin genes encoded on chromosome 1D that were thought previously to be pseudogenes. Conclusions: While breeding approaches can be used to reduce the levels of the highly immunogenic omega-5 Gliadins in wheat flour, these approaches are complicated by the genetic linkage of different classes of gluten protein genes and the finding that omega-5 Gliadins may be encoded on more than one chromosome. The work illustrates the importance of detailed knowledge about the genomic regions harboring the major gluten protein genes in individual wheat cultivars for future efforts aimed at reducing the immunogenic potential of wheat flour.
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Assessment of the Allergenic Potential of Transgenic Wheat (Triticum aestivum) with Reduced Levels of omega 5-Gliadins, the Major Sensitizing Allergen in Wheat-Dependent Exercise-Induced Anaphylaxis
Journal of Agricultural and Food Chemistry, 2015Co-Authors: Susan B. Altenbach, Roberta Lupi, Florence Pineau, Charlene K. Tanaka, Martine Drouet, Etienne Beaudouin, Martine Morisset, Sandra Denery-papiniAbstract:The omega 5-Gliadins are the major sensitizing allergens in wheat-dependent exercise-induced anaphylaxis (WDEIA). In this study, two-dimensional immunoblot analysis was used to assess the allergenic potential of two transgenic wheat lines in which omega 5-Gliadin genes were silenced by RNA interference. Sera from 7 of 11 WDEIA patients showed greatly reduced levels of immunoglobulin E (IgE) reactivity to omega 5-Gliadins in both transgenic lines. However, these sera also showed low levels of reactivity to other gluten proteins. Sera from three patients showed the greatest reactivity to proteins other than omega 5-Gliadins, either high-molecular-weight glutenin subunits (HMW-GSs), alpha-Gliadins, or non-gluten proteins. The complexity of immunological responses among these patients suggests that flour from the transgenic lines would not be suitable for individuals already diagnosed with WDEIA. However, the introduction of wheat lacking omega 5-Gliadins could reduce the number of people sensitized to these proteins and thereby decrease the overall incidence of this serious food allergy.
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Wheat Gliadins modified by deamidation are more efficient than native Gliadins in inducing a Th2 response in Balb/c mice experimentally sensitized to wheat allergens
Molecular Nutrition and Food Research, 2012Co-Authors: Pascal Gourbeyre, Colette Larré, Sandra Denery-papini, Jean-charles Gaudin, Chantal Brossard, Marie BodinierAbstract:SCOPE: Wheat gluten proteins such as Gliadins constitute major food allergens. Gluten can be modified industrially by deamidation which increases its solubility and enhances its use as a food ingredient. Sensitization to deamidated gluten has been reported to cause severe allergic reactions with anaphylaxis. The aim of this study was therefore to compare the sensitization and elicitation potentials of native (NG) and deamidated (DG) Gliadins. The reactivity pattern of mice IgE was also compared with that of DG-allergic patients.\n\nMETHODS AND RESULTS: The ability of DG to sensitize Balb/c mice using intra-peritoneal administration with aluminium hydroxide as an adjuvant, and to elicit an allergic response after a challenge, was tested in comparison with NG. Mice sensitized with DG secreted higher levels of total IgE, IL-4, Gliadin-specific IgE and IgG1 than mice sensitized with NG. By contrast, mice sensitized with NG produced higher levels of Gliadin-specific IgG2a and INFγ. After a challenge, histamine levels were higher in mice sensitised with DG.\n\nCONCLUSIONS: DG can sensitize mice much more efficiently than NG. Moreover, this mouse model of allergy to DG revealed an IgE reactivity pattern against purified Gliadins which was very similar to that of DG-allergic patients.
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A Recombinant omega-Gliadin-like D-Type Glutenin and an alpha-Gliadin from Wheat (Triticum aestivum): Two Immunoglobulin E Binding Proteins, Useful for the Diagnosis of Wheat-Dependent Allergies
Journal of Agricultural and Food Chemistry, 2012Co-Authors: Hamza Mameri, Sandra Denery-papini, C. Pecquet, I. Bouchez-mahiout, D. Choudat, N. Raison-peyron, Habib Chabane, Stephane Kerre, Yann Gohon, Pierre BriozzoAbstract:Among the wheat prolamins, D-type glutenins display a highly repetitive sequence similar to omega-Gliadins, but they contain a cysteine, that allows them to be included in the gluten macropolymers. An omega-Gliadin-like D-type glutenin, an alpha-Gliadin, and an omega 5-Gliadin-like D-type glutenin were obtained as recombinant proteins and compared using synchrotron radiation circular dichroism. This technique evidenced the strong thermostability of the omega 5-Gliadin-like protein. The IgE reactivity of recombinant proteins was evaluated using 45 sera from wheat-allergic patients. The sera from patients diagnosed with cutaneous hypersensitivity to hydrolyzed wheat proteins often reacted with the omega-Gliadin-like D-type glutenin and alpha-Gliadin, whereas the IgE reaction was less frequent after dietary sensitization. So, these two proteins could be useful to diagnose these diseases. The sera from patients with exercise-induced anaphylaxis recognized the omega 5-Gliadin-like protein as a positive control and, less frequently, the other proteins tested. Only some sera from patients with baker's asthma reacted with the proteins tested.
Enrique Mendez - One of the best experts on this subject based on the ideXlab platform.
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new strategy for the determination of Gliadins in maize or rice based foods matrix assisted laser desorption ionization time of flight mass spectrometry fractionation of Gliadins from maize or rice prolamins by acidic treatment
Journal of Mass Spectrometry, 2003Co-Authors: Alberto Hernando, Israel Valdes, Enrique MendezAbstract:A procedure for determining small quantities of Gliadins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) in gluten-free foods containing relatively large amounts of prolamin proteins from maize or rice is described. We report for the first time that Gliadins, the ethanol-soluble wheat prolamin fraction, can be quantitatively solubilized in 1.0 M acetic acid, while the corresponding ethanol-soluble maize or rice prolamin fraction remains insoluble in acetic acid. We describe a methodology for the detection of Gliadins in maize and rice foods based on a two-step procedure of extraction (60% aqueous ethanol followed by 1 M acetic acid). Subsequent MALDI-TOFMS analysis of the resulting acidic extract from these gluten-free foods clearly confirms the presence of a typical mass pattern corresponding to Gliadin components, ranging from 30 to 45 kDa. Depending on the percentages of maize or rice flours employed in the elaboration of these foods, the combined procedure enables levels of Gliadins from 100 to 400 ppm to be detected. The efficiency of this combined procedure corroborates enzyme-linked immunosorbent assay data for a large number of maize/rice gluten-free foods by means of direct visualization of the characteristic Gliadin mass pattern in maize or rice foods.
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direct identification of wheat Gliadins and related cereal prolamins by matrix assisted laser desorption ionization time of flight mass spectrometry
Journal of Mass Spectrometry, 1995Co-Authors: Enrique Mendez, Emilio Camafeita, San J Sebastian, I Valle, J Solis, F J Mayerposner, D Suckau, C Marfisi, F SorianoAbstract:Matrix-assisted UV laser desorption/ionization time-of-flight mass spectrometry (MALDI/TOF-MS) allows the detection of large proteins from highly complex protein mixtures such as those present in gluten prolamins. These proteins from wheat (Gliadins), barley (hordeins), rye (secalins) and probably oats (avenins) are known to cause the mucosal damage in coeliac disease (gluten-sensitive enteropathy). Gluten prolamins are structurally and chemically related, and their resolution into single components is extremely difficult. Preliminary results are presented based on a novel choice of analysis and identification of prolamins from unfractionated protein complex mixtures by MALDI/TOF-MS. The high resolution and sensitivity of this technique have allowed the elucidation of protonated molecular masses of most of the Gliadin, hordein, secalin and avenin components displaying a typical characteristic mass pattern : Gliadins showed mass signals corresponding to Gliadin components ranging from 31 to 55 kDa ; barley showed mass signals for 2-3 components with 32-38 kDa ; oats showed mass signals for 5-6 components with 30-35 kDa ; and rye showed mass signals for two main components of 32 and 39 kDa. Analysis by this technique of Gliadin-containing foods allows the immediate identification of the characteristic Gliadin mass pattern, consequently permitting easy identification of Gliadins in such samples. In summary, MALDI/TOF-MS seems to be a useful alternative technique for the identification of these cereal prolamins with a detection sensitivity of ∼50-100 ng total protein loaded
Eishin Morita - One of the best experts on this subject based on the ideXlab platform.
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ige antibodies to ω 5 Gliadin associate with immediate symptoms on oral wheat challenge in japanese children
Allergy, 2008Co-Authors: Komei Ito, Hiroaki Matsuo, M Futamura, Magnus P Borres, Y Takaoka, J Dahlstrom, Tatsuo Sakamoto, Akira Tanaka, Kunie Kohno, Eishin MoritaAbstract:Background: Gliadins have been implicated in immunoglobulin E (IgE)-mediated allergy to ingested wheat and ω-5-Gliadin is known to represent a major allergen in wheat-dependent exercise-induced anaphylaxis. Less known is whether ω-5-Gliadin is a clinically relevant allergen in children with immediate allergy to ingested wheat. This study investigates whether specific IgE antibodies to ω-5-Gliadin (sIgE-ω-5-Gliadin-ab) could be used as a marker for oral wheat challenge outcome in wheat-sensitized children. A secondary objective was to study whether the level of sIgE-ω-5-Gliadin was related to symptom severity in children with a positive challenge test. Methods: Serum samples from 88 children sensitized to wheat, of whom 35 underwent wheat challenge, were collected consecutively. sIgE-ω-5-Gliadin-ab was related to a physician’s diagnosis of wheat allergy and challenge symptoms. Results: The mean concentration of sIgE-ω-5-Gliadin-ab was 7.25 kUA/l in patients with wheat allergy and 1.08 kUA/l in patients with no wheat allergy (P < 0.01). sIgE-ω-5-Gliadin-ab was only detected in 12 of the non-wheat allergic children and 11 of them had a specific IgE to wheat below 1.30 kUA/l. Children reacting with severe symptoms upon challenge (n = 8) had increased levels of sIgE-ω-5-Gliadin-ab compared to children with moderate, mild or no symptoms (P < 0.001). Conclusions: The presence of sIgE-ω-5-Gliadin-ab is related to the reaction level to wheat challenge outcome in wheat-sensitized children. The sIgE-ω-5-Gliadin-ab was found to be associated with a strong convincing history of wheat allergy also in those cases when oral food challenge was avoided. The sIgE-ω-5-Gliadin-ab level may serve as a marker for clinical reactivity in wheat-sensitized individuals.
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exercise and aspirin increase levels of circulating Gliadin peptides in patients with wheat dependent exercise induced anaphylaxis
Clinical & Experimental Allergy, 2005Co-Authors: Hiroaki Matsuo, K. Morimoto, T Akaki, Sakae Kaneko, K Kusatake, T Kuroda, Hiroyuki Niihara, Michihiro Hide, Eishin MoritaAbstract:Summary Background Food-dependent exercise-induced anaphylaxis (FDEIA) is an allergic reaction characteristically induced by intense exercise combined with the ingestion of causative food. Recent reports have shown that aspirin intake is a contributing factor in some patients with FDEIA. Wheat is known to be the most frequent causative food, and the IgE-binding epitopes of a major wheat allergen (ω-5 Gliadin) in wheat-dependent exercise induced anaphylaxis (WDEIA) have already been clarified. However, the mechanism of eliciting the symptom in WDEIA remains not fully understood. Objectives The aim of this study was to examine the relationship of serum Gliadin levels and allergic symptoms induced by exercise or aspirin in patients with WDEIA. Methods Six patients with a history of recurrent anaphylaxis associated with wheat ingestion were diagnosed as having WDEIA by the provocation test, which included wheat ingestion, exercise, aspirin intake and a combination of these challenges. During the tests, serum levels of Gliadins were monitored by Gliadin-specific sandwich ELISA. The effects of exercise and aspirin on serum Gliadin levels were also investigated in four healthy subjects. Results Immunoreactive Gliadins appeared in the sera of patients during the provocation test with both wheat-exercise and wheat-aspirin challenges in parallel with allergic symptoms. Serum Gliadin levels also increased under the two same challenge conditions in the healthy subjects, although they exhibited no allergic symptoms. However, low levels of Gliadin were detected in the sera of both patients and healthy subjects when challenged with wheat alone. Conclusion We demonstrated for the first time that blood Gliadin levels correlate with clinical symptoms induced by exercise and aspirin in patients with WDEIA. These findings suggest that exercise and aspirin facilitate allergen absorption from the gastrointestinal tract.
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Fast ω-Gliadin is a major allergen in wheat-dependent exercise-induced anaphylaxis
Journal of Dermatological Science, 2003Co-Authors: Eishin Morita, Hiroaki Matsuo, Shoji Mihara, K. Morimoto, A.w.j. Savage, A.s. TathamAbstract:Abstract Background: Wheat-dependent exercise-induced anaphylaxis is an anaphylaxy induced by physical exercise after ingestion of wheat. An immediate-type hypersensitivity to water/salt-insoluble fraction of wheat proteins (gluten) has been considered to underlie in this disease. Objective: The aim of the study is to determine the major allergen in Japanese patients with wheat-dependent exercise-induced anaphylaxis by using a panel of purified wheat Gliadins and glutenins. Methods: Water/salt-insoluble wheat proteins, α-Gliadin, β-Gliadin, γ-Gliadin, fast ω-Gliadin, slow ω-Gliadin, high molecular weight glutenin and low molecular weight glutenin, were purified, and five patients with wheat-dependent exercise-induced anaphylaxis, whose diagnose had been determined by positive-challenge test, were evaluated for skin prick test, dot-blotting test and CAP–RAST inhibition test by using these purified wheat proteins. Results: The fast ω-Gliadin was the most potent allergen among these water/salt-insoluble proteins when evaluated by skin prick test and dot-blotting test. Fast and slow ω-Gliadin, and γ-Gliadin caused dose-dependent inhibition of the serum IgE-binding to solid-phase gluten in the patients. The incubation with fast ω-Gliadin of the patient's serum caused dose-dependent inhibition in the IgE-binding to γ-Gliadin as well as slow ω-Gliadin, indicating a cross-reactivity of these proteins in IgE-binding. Conclusion: We concluded that fast ω-Gliadin is a major allergen among these water/salt-insoluble proteins for wheat-dependent exercise-induced anaphylaxis in Japanese patients, and IgE against fast ω-Gliadin cross-reacts to γ-Gliadin and slow ω-Gliadin.
Herbert Wieser - One of the best experts on this subject based on the ideXlab platform.
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effect of hydrostatic pressure and temperature on the chemical and functional properties of wheat gluten studies on gluten Gliadin and glutenin
Journal of Cereal Science, 2007Co-Authors: Rolf Kieffer, F Schurer, P Kohler, Herbert WieserAbstract:Abstract The effect of hydrostatic pressure (0.1–800 MPa) in combination with various temperatures (30–80 °C) on the chemical and physical properties of wheat gluten, Gliadin and glutenin was studied. Chemical changes of proteins were determined by extraction, reversed-phase high-performance liquid chromatography (HPLC), sodium dodecylsulphate (SDS) polyacrylamide gel electrophoresis (PAGE), circular dichroism (CD) spectroscopy, thiol measurement and studies on disulphide bonds. Rheological changes were measured by extension tests and dynamic stress rheometry. Treatment of gluten with low pressure (200 MPa) and temperature (30 °C) increased the proportion of the ethanol-soluble fraction (ESF) and decreased gluten strength. The enhancement of both pressure and temperature provoked a strong reduction of the ESF and the thiol content of gluten. Within Gliadin types, cysteine containing α - and γ -Gliadins, but not cysteine-free ω -Gliadins were sensitive to pressure and were transferred to the ethanol-insoluble fraction. Disulphide peptides isolated from treated gluten confirmed that cleavage and rearrangement of disulphide bonds were involved in pressure-induced reactions. Increased pressure and temperature induced a significant strengthening of gluten, and under extreme conditions (e.g. 800 MPa, 60 °C), gluten cohesivity was lost. Isolated Gliadin and glutenin reacted differently: solubility, HPLC and SDS-PAGE patterns of Gliadin having a very low thiol content were not influenced by pressure and heat treatment; only conformational changes were detected by CD spectroscopy. In contrast, the properties of isolated glutenin having a relatively high thiol content were strongly affected by high pressure and temperature, similar to the effects on total gluten.
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cross linking to tissue transglutaminase and collagen favours Gliadin toxicity in coeliac disease
Gut, 2006Co-Authors: Walburga Dieterich, Herbert Wieser, Werner Seilmeier, Birgit Esslinger, Dagmar Trapp, Eckhart G Hahn, Thomas Huff, Detlef SchuppanAbstract:Background and aims: Intestinal inflammation in coeliac disease is driven by the gluten fraction of wheat proteins. Deamidation or cross linking of gluten peptides by tissue transglutaminase (tTG), the coeliac disease autoantigen, creates potent T cell stimulatory peptides. Therefore, our aim was to identify the reaction patterns of gluten peptides, intestinal extracellular matrix proteins, and tTG. Methods: tTG activity was analysed by incorporation of monodansyl cadaverine into Gliadins. Fluorescence labelled tTG reactive short Gliadin peptides were used to demonstrate their deamidation and explore their cross linking patterns with tTG itself or extracellular matrix proteins. Patient sera and controls were checked for autoantibodies to matrix proteins. Results: Gliadins α1–α11, γ1–γ6, ω1–ω3, and ω5 were substrates for tTG. tTG catalysed the cross linking of Gliadin peptides with interstitial collagen types I, III, and VI. Coeliac patients showed increased antibody titres against the collagens I, III, V, and VI. Conclusions: tTG formed high molecular weight complexes with all tested Gliadins. As all tested Gliadins were substrates for tTG, the tTG catalysed modifications were not restricted to single Gliadin types and epitopes. Furthermore, haptenisation and long term immobilisation of Gliadin peptides by tTG catalysed binding to abundant extracellular matrix proteins could be instrumental in the perpetuation of intestinal inflammation and some associated autoimmune diseases in coeliac disease.
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Relation between Gliadin structure and coeliac toxicity
Acta Paediatrica, 1996Co-Authors: Herbert WieserAbstract:Gliadin, the alcohol-soluble protein fraction of wheat, contains the factor toxic for coeliac patients. The numerous components of Gliadin can be classified according to their primary structure into omega 5-, omega 1,2-, alpha- and gamma-type. Both omega-types have almost entirely repetitive amino acid sequences consisting of glutamine, proline and phenylalanine. alpha- and gamma-type Gliadins contain four and five different domains, respectively, and are homologous within the domains III and V. Unique for each alpha- and gamma-type is domain I, which consists mostly of repetitive sequences rich in glutamine, proline and aromatic amino acids. Coeliac toxicity of Gliadin is not destroyed by digestion with gastropancreatic enzymes. In vivo testing established the toxicity of alpha-type Gliadins and in vitro testing of Gliadin peptides revealed that domain I of alpha-type Gliadins is involved in activating coeliac disease. The sequences -Pro-Ser-Gln-Gln- and -Gln-Gln-Gln-Pro- were demonstrated to be common for toxic Gliadin peptides. Most of the in vivo and in vitro studies of synthetic peptides confirmed the importance of one or both of these sequences. Cultivated hexaploid, tetraploid and diploid wheat species do not differ significantly in potential toxic sequences of alpha-type Gliadins.
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The location of disulphide bonds in α-type Gliadins
Journal of Cereal Science, 1995Co-Authors: S. Müller, Herbert WieserAbstract:Abstract Gliadin prepared from gluten of the cultivar Rektor by extraction with 70% (v/v) aqueous ethanol adjusted to pH 5.5 was separated by RP-HPLC. Amongst 23 components obtained, two α-type Gliadins (α3- and α8-Gliadin) were selected for the determination of disulphide bonds. After both proteins were digested with thermolysin, differential RP-HPLC (chromatography prior to and after reduction of disulphide bonds) was used for the detection of cystine peptides. Two cystine peptides from α3-Gliadin and three cystine peptides from α8-Gliadin were isolated by RP-HPLC. The resulting peptides were reduced and alkylated with 4-vinylpyridine, separated by RP-HPLC and their amino acid sequences determined. The cystine peptides from both α-type Gliadins had similar structures, and the corresponding fragments had homologous sequences. One cystine peptide of each Gliadin was composed of three fragments linked by two disulphide bonds. The second cystine peptide consisted of two fragments linked by one disulphide bond. The third cystine peptide derived from α8-Gliadin was different from the second peptide in one position of the sequences (glutamic acid instead of glutamine). Comparing complete sequences of α-type Gliadins described in the literature, the cystine peptides from α3- and α8-Gliadins were identical with corresponding sequences of clones A1235 and A212, respectively 11 . The structures of the cystine peptides analysed indicate one intramolecular disulphide bond within domain III of α-type Gliadins and two disulphide bonds between domains III and V. The linkages found correspond to homologous linkages determined for low M r subunits of glutenin and glutenin-bound γ-type Gliadins 6 . Obviously, these intramolecular disulphide bonds are not linked randomly, but are strongly directed.
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Quantitative Determination of Gliadin Subgroups from Different Wheat Cultivars
Journal of Cereal Science, 1994Co-Authors: Herbert Wieser, Werner Seilmeier, H. D. BelitzAbstract:Abstract Sixteen wheat cultivars originating from different countries and genotypes and characterised by different technological properties were analysed for the amount and proportion of ω- and α-type and γ-type Gliadin subgroups. The optimal extraction of Gliadin from flour was achieved with 60% (v/v) aqueous ethanol after pre-extraction with a salt solution. The separation and quantitative determination was performed by reversed-phase high-performance liquid chromatography on C18 silica gel. The amount of both total Gliadin and subgroups revealed distinct inter-cultivar differences. Within the subgroups, the α-type Gliadins were generally present in greatest amount, followed by the γ-type Gliadins, whereas the ω-Gliadins were present at lower levels. A strong statistical relationship was found between the protein content of the flours and the amounts of total Gliadin, α-type and γ-type Gliadins. Other flour properties (baking volume, SDS-sedimentation volume, dough resistance and extensibility) were correlated only weakly with Gliadin contents, with only ω1,2-Gliadins and γA-Gliadins showing moderate negative and positive effects, respectively. The proportions of ω- and α-type and γ-type Gliadins found within the total Gliadin fraction covered broad ranges (α: 43·9-59·9%, γ: 30·5-45·6%, ω: 6·2-20·0%). The proportions of the Gliadin subgroups were to some extent related to the genotype of the cultivar. Thus, high proportions of ω-Gliadins (17-20%) were typical for wheat/rye hybrids. In view of the inter-cultivar variation observed, the value of immunochemical assays developed for the analysis of total Gliadin in food is restricted if the reactivity of the antibodies used is not directed to all types of Gliadin components but to the minor subgroups of ω-Gliadins.
Susan B. Altenbach - One of the best experts on this subject based on the ideXlab platform.
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Towards reducing the immunogenic potential of wheat flour: omega Gliadins encoded by the D genome of hexaploid wheat may also harbor epitopes for the serious food allergy WDEIA
BMC Plant Biology, 2018Co-Authors: Susan B. Altenbach, Sandra Denery-papini, You-ran Jang, Annamaria Simon-buss, Sun-hyung Lim, Naxin Huo, Han-chang Chang, Florence Pineau, Chon-sik Kang, Jong-yeol LeeAbstract:Background: Omega-5 Gliadins are a group of highly repetitive gluten proteins in wheat flour encoded on the 1B chromosome of hexaploid wheat. These proteins are the major sensitizing allergens in a severe form of food allergy called wheat-dependent exercise-induced anaphylaxis (WDEIA). The elimination of omega-5 Gliadins from wheat flour through biotechnology or breeding approaches could reduce the immunogenic potential and adverse health effects of the flour. Results: A mutant line missing low-molecular weight glutenin subunits encoded at the Glu-B3 locus was selected previously from a doubled haploid population generated from two Korean wheat cultivars. Analysis of flour from the mutant line by 2-dimensional gel electrophoresis coupled with tandem mass spectrometry revealed that the omega-5 Gliadins and several gamma Gliadins encoded by the closely linked Gli-B1 locus were also missing as a result of a deletion of at least 5.8Mb of chromosome 1B. Two-dimensional immunoblot analysis of flour proteins using sera from WDEIA patients showed reduced IgE reactivity in the mutant relative to the parental lines due to the absence of the major omega-5 Gliadins. However, two minor proteins showed strong reactivity to patient sera in both the parental and the mutant lines and also reacted with a monoclonal antibody against omega-5 Gliadin. Analysis of the two minor reactive proteins by mass spectrometry revealed that both proteins correspond to omega-5 Gliadin genes encoded on chromosome 1D that were thought previously to be pseudogenes. Conclusions: While breeding approaches can be used to reduce the levels of the highly immunogenic omega-5 Gliadins in wheat flour, these approaches are complicated by the genetic linkage of different classes of gluten protein genes and the finding that omega-5 Gliadins may be encoded on more than one chromosome. The work illustrates the importance of detailed knowledge about the genomic regions harboring the major gluten protein genes in individual wheat cultivars for future efforts aimed at reducing the immunogenic potential of wheat flour.
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Assessment of the Allergenic Potential of Transgenic Wheat (Triticum aestivum) with Reduced Levels of omega 5-Gliadins, the Major Sensitizing Allergen in Wheat-Dependent Exercise-Induced Anaphylaxis
Journal of Agricultural and Food Chemistry, 2015Co-Authors: Susan B. Altenbach, Roberta Lupi, Florence Pineau, Charlene K. Tanaka, Martine Drouet, Etienne Beaudouin, Martine Morisset, Sandra Denery-papiniAbstract:The omega 5-Gliadins are the major sensitizing allergens in wheat-dependent exercise-induced anaphylaxis (WDEIA). In this study, two-dimensional immunoblot analysis was used to assess the allergenic potential of two transgenic wheat lines in which omega 5-Gliadin genes were silenced by RNA interference. Sera from 7 of 11 WDEIA patients showed greatly reduced levels of immunoglobulin E (IgE) reactivity to omega 5-Gliadins in both transgenic lines. However, these sera also showed low levels of reactivity to other gluten proteins. Sera from three patients showed the greatest reactivity to proteins other than omega 5-Gliadins, either high-molecular-weight glutenin subunits (HMW-GSs), alpha-Gliadins, or non-gluten proteins. The complexity of immunological responses among these patients suggests that flour from the transgenic lines would not be suitable for individuals already diagnosed with WDEIA. However, the introduction of wheat lacking omega 5-Gliadins could reduce the number of people sensitized to these proteins and thereby decrease the overall incidence of this serious food allergy.
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comparative proteomic analysis of the effect of temperature and fertilizer on Gliadin and glutenin accumulation in the developing endosperm and flour from triticum aestivum l cv butte 86
Proteome Science, 2013Co-Authors: William J Hurkman, Charlene K. Tanaka, William H. Vensel, Roger Thilmony, Susan B. AltenbachAbstract:Background Flour quality is largely determined by the gluten proteins, a complex mixture of proteins consisting of high molecular weight-glutenin subunits (HMW-GS), low molecular weight-glutenin subunits (LMW-GS), and α-, γ-, and ω-Gliadins. Detailed proteomic analyses of the effects of fertilizer and high temperature on individual Gliadin and glutenin protein levels are needed to determine how these environmental factors influence flour quality.
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analysis of expressed sequence tags from a single wheat cultivar facilitates interpretation of tandem mass spectrometry data and discrimination of gamma Gliadin proteins that may play different functional roles in flour
BMC Plant Biology, 2010Co-Authors: Susan B. Altenbach, William H. Vensel, Frances M DupontAbstract:The gamma Gliadins are a complex group of proteins that together with other gluten proteins determine the functional properties of wheat flour. The proteins have unusually high levels of glutamine and proline and contain large regions of repetitive sequences. While most gamma Gliadins are monomeric proteins containing eight conserved cysteine residues, some contain an additional cysteine residue that enables them to be linked with other gluten proteins into large polymers that are critical for flour quality. The ability to differentiate among the gamma Gliadins is important for studies of wheat flour quality because proteins with similar sequences can have different effects on functional properties. The complement of gamma Gliadin genes expressed in the wheat cultivar Butte 86 was evaluated by analyzing publicly available expressed sequence tag (EST) data. Eleven contigs were assembled from 153 Butte 86 ESTs. Nine of the contigs encoded full-length proteins and four of the proteins contained nine cysteine residues. Only one of the encoded proteins was a perfect match with a sequence reported in NCBI. Contigs from four different publicly available EST assemblies encoded proteins that were perfect matches with some, but not all, of the Butte 86 gamma Gliadins and the complement of identical proteins was different for each assembly. A specialized database that included the sequences of Butte 86 gamma Gliadins was constructed for identification of flour proteins by tandem mass spectrometry (MS/MS). In a pilot experiment, proteins corresponding to six Butte 86 gamma Gliadin contigs were distinguished by MS/MS, including one containing the extra cysteine residue. Two other proteins were identified as one of two closely related Butte 86 proteins but could not be distinguished unequivocally. Unique peptide tags specific for Butte 86 gamma Gliadins are reported. Inclusion of cultivar-specific gamma Gliadin sequences in databases maximizes the number and quality of peptide identifications and increases sequence coverage of these gamma Gliadins by MS/MS. This approach makes it possible to distinguish closely related proteins, to associate individual proteins with sequences of specific genes, and to evaluate proteomic data in a biological context to better address questions about wheat flour quality.